As a therapeutic method for regenerating adipose tissues, there is a method for using a therapeutic agent comprising stem cells, which are cultured with a matrix having an adipose tissue three-dimensionally cultured in a hydrogel. After three-dimensionally culturing the tissue in a hydrogel, a growth factor and an extracellular matrix may be secreted from the stem cells. However, to use the therapeutic agent as an injectable preparation type, it was inconvenient to remove the hydrogel in the form of a film from a culture container and to treat the tissue with a lyase. To overcome such inconvenience, a therapeutic method for regenerating tissues by autologous fat grafting or directly transplanting stem cells has been developed.
In the case of autologous fat grafting, the method uses parts of the body of a subject under operation. Therefore, the method incurs no issue of tissue or immune rejection and no observed immune response. However, the adipose tissue is highly oxygen-dependent and interacts with neighboring cells while having many blood vessels around. The grafted fat hardly exhibits a blood vessel-forming ability, and thus has disadvantages. For example, a cell apoptosis or cell necrosis may be induced due to hypoxia, and the subject may need to receive several procedures as the rate of engraftment is not high.
Formerly, stem cells have frequently been used to restore damaged tissues that have been limited by surgery or drug therapy. For example, biopolymers such as hyaluronic acid and collagen are used for stem cell transplantation. Since stem cells can differentiate into various cells including adipocytes, these stem cells have a wide range of applications. However, since the survival rate and the engraftment rate are low once they are placed into the body, the efficiency is reduced. Also, there is a risk that undifferentiated stem cells can form tumors.
Currently, a method for differentiating stem cells into adipocytes for tissue regeneration generally includes treating differentiation-inducing materials such as insulin, dexamethasone, isobutylmethylxanthine, etc. on stem cells and culturing them for a long time. However, the above-mentioned stem cell differentiation-inducing materials are expensive and are not effective for differentiation only by a single component. Thus, they have disadvantages. For example, they must be treated by mixing various substances, and the efficiency of cell differentiation is low. This is problematic.
On the other hand, conventionally, attempts have been made to use a culture solution obtained by culturing stem cells as a cosmetic. In general, a culture medium containing an appropriate amount of antibiotics and serum is used for culturing stem cells. Most of the stem cell culture solutions developed as cosmetic compositions use a normal culture medium, and a cosmetic composition including a liposome in which a culture solution of stem cells is encapsulated in a liposome. Further, a cosmetic composition using a culture medium prepared without the ingredients that are not permitted as raw materials for cosmetics and a cosmetic composition containing a serum-free culture medium, and the like have been developed.
The culture media are substances containing proteins, amino acids, hormones and growth factors for cell proliferation. The media have been prepared in a very sophisticated manner and supplied. However, since the cell culture media, antibiotics and serum have risks that are not proven as safe, they should be used only for research purposes and their use for the human body is prohibited. The components included in the culture media, such as choline chloride, hypoxanthine-sodium salt, thymidine, putrescine dihydrochloride, ferric nitrate, L-glutamine and the like, are not permitted as raw materials for cosmetics. Thus, the use of such culture media is not suitable for a cosmetic composition. As such, the culture media contain various proteins, cytokines, growth factors and the like secreted by stem cells. In contrast, they also contain components such as waste products secreted as cells grow, antibiotics added to prevent contamination, or animal-derived serum, etc. Thus, they are highly likely to pose various risks when used on the skin.
The components of the stem cell culture solutions to be used for cosmetics are limited. Also, during the process of encapsulating into the liposomes, the deterioration and contamination of the components of the culture solutions, and an additional treatment process of encapsulating with liposomes are required. Thus, the technique of encapsulating the stem cell culture solutions with liposomes including lipids to increase the skin absorption rate of the culture solutions is also limited in the use for a cosmetic.
To complement the disadvantages of these stem cell culture solutions, techniques for using stem cell-derived exosomes have been developed. Stem cells are usually cultured in a medium containing antibiotics and serum. Bio-nanoparticles secreted from various cells present in multicellular organisms including humans can be classified into exosomes and micro-vesicles depending on their size and difference in secretion mechanism. It is known that exosomes, which are vesicles of membrane structures secreted from various types of cells, play a variety of roles. For example the roles include transferring membrane components, proteins, RNA, etc. by binding to other cells and tissues. Most of secretomes including the exosomes are obtained from a cell culture supernatant. Thus, under a stem cell-derived exosome isolation method currently used, it is difficult to completely purify the exosomes due to interference by proteins in the medium or serum in the step of isolating the secretomes including the exosomes.
Accordingly, the present inventors have isolated exosomes from stem cells and discovered that the stem cell-derived exosomes have the effects of stem cell differentiation, adipose tissue regeneration, whitening, wrinkle improvement and skin regeneration.